RecQ proteins are a conserved, ubiquitous class of DNA helicases essential for genomic stability. Heritable mutations in RecQ Werner syndrome protein (WRN) or RecQ Bloom syndrome protein (BLM) cause remarkably elevated cancer predispositions and WRN mutations cause an additional aging phenotype. WRN and BLM have divergent roles; yet, both may act as tumor suppressor proteins and maintain genetic integrity by functioning with DNA replication and double-strand break repair proteins. Integrated structural, mutational, and biochemical analyses in the Yannone and Tainer labs will address several unanswered questions as to how WRN and BLM act in preserving genome integrity. We aim to define at the molecular level how WRN and BLM detect and bind DNA substrate, open and unwind target DNA, cleave and resolve DNA intermediates, and coordinate initial protein hand-offs to aid DNA replication and repair pathways. To accomplish this and test specific hypotheses on the molecular basis for WRN and BLM activities, we will biochemically and structurally characterize WRN and BLM components plus their complexes with DNA and key protein partners. Systematic structural and mutational analyses will define functional conformational states and interfaces and suggest mutations to uncouple DNA and/or protein binding events that modulate WRN and BLM activities. We will define the consequences of WRN and BLM conformations and interactions by protein structural studies, biochemical assays in vitro, and functional in vivo assays with our collaborators. These proposed experiments will create a molecular picture of the functional states, protein-DNA, and protein-protein interactions that orchestrate WRN- and BLM-mediated DNA metabolism. Taken together, the anticipated results will provide a detailed molecular understanding of processes underlying the aging and cancer predispositions associated with WRN and BLM helicase deficiencies and create a paradigm for RecQ helicase activities in choreographing replication, recombination and repair events to avoid cancer and aging.